The ability to control Ad tropism in vivo to restrict infection to cancer cells will certainly increase the safety and efficacy of virus-mediated treatment while expanding the use of Ad vectors beyond loco-regional administration in humans. The achievement of such targeted infection requires both ablation of broad viral tropism and engineering of a novel mechanism of selective cancer cell recognition. In this respect, tropism of widely used Ad of serotype 5 (Ad5) is mediated by interaction of trimeric fiber protein withcoxsackie- adenovirus receptor (CAR) and heparan sulfate proteoglycans (HSPG) as well as by binding of the Arg- Gly-Asp (RGD) motif in penton to aV-integrins on the cell surface. Therefore, elimination of CAR, HSPG, and integrin recognition coupled with virus binding to a tumor-specific receptor would represent a valid means to ablate an endogenous viral tropism while achieving retargeting of Ad vector to cance* cells. We hypothesize that the genetic incorporation of high fidelity ligand derived from single-chain antibody (scFv) with defined specificity for tumor antigen into the viral capsid would confer Ad targeting capability to cancer cells. In support of our hypothesis, we have demonstrated the feasibility of employment of minor capsid protein IX (pIX) as a novel locale for incorporation of ligands of augmented size/complexity, thus, offering improved targeting potentials compared with other sites in penton and hexon proteins investigated for this purpose. In an effort to design tumor-targeted Ad vector we have demonstrated our ability to engineer C6.5 scFv into pIX in a manner commensurate with virion integrity and binding to the c-erbB-2 oncoprotein, a molecule of established therapeutic relevance in breast and ovarian cancer. In order to achieve ablation of broad native viral tropism, which is at cross-purposes with our goal of precise and absolute targeting to cancer cells, we propose to combine the replacement of Ad5 fiber for the short Ad41 fiber devoid of all known determinant of Ad tropism with the RGD motif deletion in penton base. The purpose of this proposal is to design an oncotropic Ad that embodies genetic virion modifications providing tumor-targeting capability while eliminating ectopic infection of normal tissues in the context of systemic vascular vector administration.